Transcription - Lecture 2

Question 1

Briefly describe the structure of a prokaryotic gene.

Answer 1

contains promoter region, RNA coding region, and terminator region

Question 2

Describe the RNA polymerase of prokaryotes.

Answer 2

consists of a core haloenzyme: ⍺2ββ’⍵σ

Question 3

Describe all the subunits of the prokaryotic RNA polymerase.

Answer 3

⍺: involved in the assembly of the tetrameric core
β: contains rNTP binding site
β’: contains DNA template binding region
⍵: stabilizes core
σ: gives the RNA polymerase specificity for the gene

Question 4

What is the -35 sequence in prokaryotes?

Answer 4

5’ TTGACA 3’

Question 5

What is the -10 sequence in prokaryotes?

Answer 5

5’TATAAT3’

Question 6

Describe the initiation of transcription in prokaryotes.

Answer 6

• Sigma factor recognizes and binds to the -35 element in the promoter region
• Transcription initiates about 5-9 base pairs down from the end of the -10 sequence, which is the 5’ end of a purine (+1 position)
• The RNA polymerase will recognize this purine and start transcribing the 3’ to 5’ DNA template strand

Question 7

Describe elongation of transcription in prokaryotes.

Answer 7

• Sigma factor is release and RNA polymerase begins to move along the 3’ to 5’ DNA template strand
• Transcription bubble occurs as RNA poly moves along strand
• RNA polymerase has helicase that assists in unwinding
• Topoisomerase removes positive supercoils

Question 8

Describe termination of transcription in prokaryotes.

Answer 8

• At the end of prokaryotic genes there is a palindromic sequence
• When these genes are transcribed, they form a stem loop through H-bonds
• When the hairpin forms, the polymerase pauses
• The area downstream from the hairpin is a U:A rich region so it weakly hydrogen bonded which allows for the hairpin structure to be released

Question 9

What does RNA polymerase I,II, and II each transcribe?

Answer 9

I - large rRNAs
II - Pre-mRNA, snRNAs, snoRNAs, and miRNAs
III - tRNAs, small rRNAs, snRNAs, and miRNAs

Question 10

What is the sequence of the -25 box in eukaryotes?

Answer 10

5’ TATAAA 3’

Question 11

Describe the initiation of transcription in eukaryotes.

Answer 11

• Involves ordered assembly of TFII A,B,D,E,F, and H
• TFIID assembles first at the TATA box followed by the remaining general transcription factors and Pol II
• This forms preinitiation complex (PIC)

Question 12

Describe termination of transcription in eukaryotes.

Answer 12

• RNA poly II transcribes past coding sequence of gene
• RNA poly II continues transcribing while mRNA is cleaved at a sequence located by Rat1
• Rat1 degrades the un-cleaved mRNA until it reaches the RNA poly and transcription stops

Question 13

What is ⍺-amanitin?

Answer 13

an inhibitor of the initiation and elongation steps of eukaryotic transcription

Question 14

What is the difference between the -35 sequence in prokaryotes and eukaryotes?

Answer 14

prokaryotes: 5’TTGACA3’, sigma factor recognition site
eukaryotes: GC box, binding site for 2nd transcription factor

Question 15

What is the functioning of the -10 sequence in prokaryotes?

Answer 15

to aid in unwinding